SUMMARYApproximately 50% of Wistar "BB" rats spontaneously develop overt diabetes mellitus characterized by loss of /3-cells and "insulitis." To define abnormalities of immunoregulation in these rats, we quantitated their major circulating lymphocyte subsets. Independent of the development of diabetes, we found the BB rats to have a markedly increased percentage of circulating B lymphocytes which is secondary to a severe T-cell lymphocytopenia, with the major circulating T-cell subset reacting with monoclonal antibody W3/25 markedly decreased. This lymphocytopenia is present in every animal studied and contrasted with studies of the nondiabetic Wistar strain from which the "BB" rats were developed. DIABETES 30:887-889, October 1981. T he spontaneously diabetic Wistar rat (the "BB rat") has become an important animal model of type I insulin-dependent diabetes mellitus. -2Approximately 50% of these nonobese outbred Wistar rats between 60 and 120 days of age develop severe hyperglycemia associated with /3-cell depletion, insulitis, and ketoacidosis. Of major importance is the finding of Like et al. that various immunosuppressive regimens (including administration of anti-lymphocyte globulin) can prevent the onset of overt diabetes in these animal.3 -4 It is postulated that the development of diabetes mellitus is secondary to inherited abnormalities of immunologic function. In this report we describe our findings that all BB rats, independent of the development of overt diabetes, have a specific deficiency of T-cells reacting with monoclonal antibody W3/25. METHODSNineteen rats of the BB strain and 20 rats of the control, nondiabetic Wistar strain (male and female) were kindly provided by Dr. P. Thibert. Rats were screened for glucosuria using Testape (Eli Lilly and Company, Indianapolis, Indiana) approximately every 3 days, and plasma glucose was measured using a Beckman glucose analyzer (Beckman Instruments, Fullerton, California) at least once a week. The onset of diabetes was abrupt in the six animals that became diabetic. Plasma glucose at the time of diagnosis of diabetes ranged from 284 to 649 mg/dl. Approximately every 2 wk, animals were gently anesthetized with ether, and 1 ml of blood was withdrawn from a tail vein into heparinized tubes for determination of white blood count, differential and lymphocyte subsets. White blood count was determined using a Coulter Zf counter and differential using Wright's stained slides. Peripheral white blood cells were then applied to a Ficoll-Hypaque gradient (24:10 v,v; 9% Ficoll 400:34% Hypaque M) and the lymphocytes and monocytes present at the interface of the gradient were harvested. This lymphocyte fraction was then incubated at 37°C for 45 min to remove cytophilic immunoglobulin, and then reacted with rhodamine conjugated F (ab') 2 anti-rat antibody (Cappel) at 4°C for 45 min, followed by three washes with phosphatebuffered saline containing 1% bovine albumin. To determine the percentage of B lymphocytes, the number of cells with rim fluorescence was determined usi...